Dynamic Activity of miR-125b and miR-93 during Murine Neural Stem Cell Differentiation In Vitro and in the Subventricular Zone Neurogenic Niche

Annalisa Lattanzi,Pieter Mestdagh,Daniela Corno,Tiziano Di Tomaso,Angela Gritti,Frank Speleman,Luigi Naldini,Bernhard Gentner,Cesar V Borlongan

doi:10.1371/journal.pone.0067411

Abstract

Several microRNAs (miRNAs) that are either specifically enriched or highly expressed in neurons and glia have been described, but the identification of miRNAs modulating neural stem cell (NSC) biology remains elusive. In this study, we exploited high throughput miRNA expression profiling to identify candidate miRNAs enriched in NSC/early progenitors derived from the murine subventricular zone (SVZ). Then, we used lentiviral miRNA sensor vectors (LV.miRT) to monitor the activity of shortlisted miRNAs with cellular and temporal resolution during NSC differentiation, taking advantage of in vitro and in vivo models that recapitulate physiological neurogenesis and gliogenesis and using known neuronal- and glial-specific miRNAs as reference. The LV.miRT platform allowed us monitoring endogenous miRNA activity in low represented cell populations within a bulk culture or within the complexity of CNS tissue, with high sensitivity and specificity. In this way we validated and extended previous results on the neuronal-specific miR-124 and the astroglial-specific miR-23a. Importantly, we describe for the first time a cell type- and differentiation stage-specific modulation of miR-93 and miR-125b in SVZ-derived NSC cultures and in the SVZ neurogenic niche in vivo, suggesting key roles of these miRNAs in regulating NSC function.

Highlights

MicroRNAs are small non-coding-RNAs that regulate the expression of a large fraction of mRNA in a sequencespecific manner [1] through translational repression and/or transcript destabilization [2], [3]
Data were analyzed by one-way analysis of variance followed by Bonferroni’s posttest. *** p,0.001 (NeuN), ˆp,0.01 (GFAP) versus neuronal nuclear antigen (NeuN) and glial fibrillary protein (GFAP) values of LV.CTRL-injected mice. (F, G) Quantification and representative images of miR-125b activity after striatal injection of bdLV.miRT125b
We identified miR-125b and miR-93 as abundantly expressed in subventricular zone (SVZ) neural stem/ progenitor cells, and extended our understanding on their potential involvement in the regulation of neural stem cell (NSC) function

Summary

Introduction

MicroRNAs (miRNAs) are small non-coding-RNAs that regulate the expression of a large fraction of mRNA in a sequencespecific manner [1] through translational repression and/or transcript destabilization [2], [3]. Functional studies of individual miRNAs in the CNS require techniques allowing to simultaneously monitor spatio-temporal expression patterns and cellular localization. To this end, strategies based on the visualization of miRNA-regulated reporters have been developed [8] that could overcome the low sensitivity of histology and in situ RNA expression [19], [20]. A similar strategy could possibly be used to enrich for NSCs or committed progenitors, providing large amounts of neural cells suitable for transplantation in different neurodegenerative pathologies In this perspective, a comprehensive knowledge of the modulation of PLOS ONE | www.plosone.org miRNAs in Somatic Neural Stem Cells specific miRNAs during NSC maintenance/differentiation is needed

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Discussion

Conclusion